摘要
目的:探讨METTL14介导的m^(6)A甲基化修饰Snail-1蛋白在食管鳞状细胞癌(ESCC)侵袭和上皮间质转化(EMT)中的影响和作用机制。方法:通过GEPIA分析ESCC患者METTL14的表达情况及对其生存期进行评估;实验分为si-NC组和si-METTL14组;qPCR检测METTL14基因的表达;Western blot检测Snail-1和EMT相关蛋白(E-cadherin、N-cadherin、Vimentin)的表达;Transwell实验检测细胞侵袭能力;MeRIP-qPCR检测m^(6)A修饰Snail-1的水平;加入放线菌素D(actinomycin D)检测Snail-1 mRNA稳定性。结果:ESCC患者和细胞系中METTL14的表达显著升高(P<0.01);METTL14高表达患者的生存期明显缩短(P<0.01)。si-METTL14组中METTL14 mRNA和蛋白的表达水平都显著降低(P<0.01);E-cadherin表达显著上升(P<0.01),而N-cadherin和Vimentin的表达显著降低(P<0.01);细胞侵袭个数明显减少(P<0.01)。进一步发现,si-METTL14转染显著下调Snail-1的mRNA和蛋白水平(P<0.01),且显著降低m^(6)A Snail-1水平(P<0.01)以及Snail-1 mRNA的稳定性(P<0.01)。Snail-1拯救实验证明与si-METTL14+OE-NC组相比,si-METTL14+OE-Snail-1组中E-cadherin的表达显著降低(P<0.01),N-cadherin和Vimentin的表达显著升高(P<0.01)。结论:在ESCC细胞中,METTL14和Snail-1的表达均上调;敲低METTL14可显著降低Snail-1 mRNA的稳定性,而减少其表达,从而抑制ESCC细胞侵袭和EMT过程。
Objective:To investigate the effect and mechanism of METTL14-mediated m^(6)A methylation modifying Snail-1 protein in esophageal squamous cell carcinoma(ESCC)invasion and epithelial mesenchymal transformation(EMT).Methods:GEPIA was used to analyze the expression of METTL14 in ESCC patients and to evaluate their survival.The experiment was divided into si-NC group and si-METTL14 group.The expression of METTL14 gene was detected by qPCR.The expressions of Snail-1 and EMT-related proteins(E-cadherin,N-cadherin and Vimentin)were detected by Western blot.Transwell test was used to detect cell invasion ability.The level of m^(6)A modified Snail-1 was detected by MeRIP-qPCR.The stability of Snail-1 mRNA was detected by adding actinomycin D.Results:The expression of METTL14 was significantly increased in ESCC patients and cell lines(P<0.01).Overall survival was significantly shortened in patients with high expression of METTL14(P<0.01).The mRNA and protein expression levels of METTL14 were significantly decreased in si-METTL14 group(P<0.01).E-cadherin expression was significantly increased(P<0.01),while the expressions of N-cadherin and Vimentin were significantly decreased(P<0.01).The number of cell migration was significantly decreased(P<0.01).Further,si-METTL14 transfection significantly down-regulated Snail-1 mRNA and protein levels(P<0.01),and significantly reduced m^(6)A Snail-1 levels(P<0.01)and Snail-1 mRNA stability(P<0.01).Snail-1 rescue experiment showed that the expression of E-cadherin in si-METTL14+OE-Snail-1 group significantly decreased compared with that in si-METTL14+OE-NC group(P<0.01),the expressions of N-cadherin and Vimentin were significantly increased(P<0.01).Conclusion:The expressions of METTL14 and Snail-1 were up-regulated in ESCC cells.METTL14 knockdown significantly reduced the stability of Snail-1 mRNA and reduced its expression,thus inhibiting ESCC cell invasion and EMT process.
作者
钟守平
李广顺
ZHONG Shouping;LI Guangshun(Cardiothoracic Surgery,Xi'an Central Hospital,Shaanxi Xi'an 710004,China)
出处
《现代肿瘤医学》
CAS
北大核心
2023年第19期3565-3571,共7页
Journal of Modern Oncology
